MessageAmp™ II-96 aRNA Amplification Kit is the first kit designed for high throughput amplification of mRNA and is based on the patented Van Gelder-Eberwine linear RNA amplification method. This kit takes advantage of extensive improvements incorporated into the MessageAmp™ II aRNA Kit combined with a new magnetic bead-based purification of both the cDNA and the aRNA.

Figure 3. mRNA Profiles of Mouse Tissues Stored in RNAlater. Various mouse tissues were stored in RNAlater for 1 or 4 weeks at 4°C. RNA was isolated from each tissue and analyzed using the RPA III™ Kit. 10 µg of total RNA was hybridized with 5 x 104 cpm of each of 5 combined antisense RNA probes, digested with RNase and precipitated. Products were assessed on a 5% polyacrylamide / 8 M urea gel and exposed to film for 4 hours at -80°C with an intensifying screen.

Figure 1. Robotic or Manual RNA Amplification using MessageAmp™II-96 aRNA Amplification Kit. Twelve replicates of HeLa cell total RNA inputs (5, 1, 0.3, and 0.1 µg) were processed according to the MessageAmp II-96 protocol either on the Biomek® 2000 (Robot, Beckman-Coulter) or manually (human). In vitro transcription (IVT) times of (A) 14 hours and (B) 4 hours were used. Following aRNA clean-up, yield was assessed by measuring UV absorbance at 260 nm.

Figure 3. mRNA Profiles of Mouse Tissues Stored in RNAlater. Various mouse tissues were stored in RNAlater for 1 or 4 weeks at 4°C. RNA was isolated from each tissue and analyzed using the RPA III™ Kit. 10 µg of total RNA was hybridized with 5 x 104 cpm of each of 5 combined antisense RNA probes, digested with RNase and precipitated. Products were assessed on a 5% polyacrylamide / 8 M urea gel and exposed to film for 4 hours at -80°C with an intensifying screen.

Magnetic beads are used for both the cDNA and aRNA purification steps, eliminating the need for vacuum filtration or centrifugation. The use of beads also enables elution into smaller volumes, thus providing a higher concentration of aRNA. The entire process can be performed either manually or fully automated using a robot integrated with a thermal cycler and orbital shaker. Each kit contains sufficient reagents to amplify 100 samples, thus providing four samples for pre-testing prior to using a 96 well format.